A goal of clinical research is to
deliver the right drug protocol to the right patient - based on
the right diagnosis of the disease - the underlying biology.
DLBCL progress has been made ... but we have gotten ahead of
immunohistochemistry (IHC) tests on biopsy samples do not
appear to be reproducible (reliable) and therefore are not yet
useful for guiding the approach to initial therapy for DLBCL.
Further, we are not yet at a point where a reliable finding of
cell of origin (COO) can guide the approach to therapy.
Within, Perry et al. describe the limitations of IHC tests
relative to tests using Gene Expression Profiling (GEP) on snap
frozen fresh tissue ... and the need for a reliable test that is
practical to use in the clinical setting.
The second report describes a
promising new assay that could help to select patients for
clinical trials based on the cell of origin and treatment
agents that may be more active in different cell types.
Biological Prognostic Markers in Diffuse Large B-Cell
Anamarija M. Perry, MD, Zdravko Mitrovic, MD, and Wing C.
prognostic markers have been proposed in patients with
diffuse large B-cell lymphoma (DLBCL), and the significance
of many of these that were studied before the rituximab era
need to be reassessed. Prognostic markers are assayed by a
variety of methods, most commonly by morphology and
Although widely available and relatively cheap, the
immunohistochemistry method suffers from poor
difficulty in quantification due to differences in
tissue processing, staining protocols, and inter-observer
Molecular methods such as gene expression profiling (GEP),
high-resolution array comparative genomic hybridization, and
next-generation sequencing hold great promise in elucidating
the pathogenesis and prognosis of DLBCL, but these methods
are not widely available due to substantial cost, technical
complexity, and requirement for fresh and frozen tissue.
However, more focused assays can be designed for further
studies, and the ability to apply these assays to
formalin-fixed, paraffin-embedded tissue would allow the
inclusion of large patient cohorts to improve statistical
Next-generation sequencing has detected numerous mutations
in patients with DLBCL, but whether any of those mutations
are important for prognosis, alone or in combination, is not
yet answered. However, such global studies allow us to
examine markers in the context of other modifying factors
and hence overcome the problems of single-marker studies.
For example, BCL2 may be an important biomarker, but its
clinical significance is influenced by other factors such as
other biological activities of the pathway that leads to its
expression, the coexisting factors such as MYC
translocation, and possibly even mutations that may affect
its biological activities. Thus, studying BCL2 expression
alone as a biomarker may not generate reproducible results
from different populations.
A more global approach may allow all of these factors to be
included in the analysis, thereby producing a more
meaningful biomarker profile.
At the present time, there is no consensus on which
biological prognostic markers should be routinely assessed
in patients with DLBCL, and practices vary widely among
different institutions. With more global approaches, such as
those noted above, the ability to assess biomarkers in the
cellular or tumor context may be possible, resulting in a
better understanding of their biological and prognostic
A reliable test for cell of origin in DLBCL appears to be on
ASH 2013: Determining Cell-Of-Origin
Subtypes In DLB Lymphoma Using Gene Expression Profiling On
Formalin-Fixed Paraffin-Embedded Tissue – An L.L.M.P.P.
The diffuse large B-cell
lymphoma (DLBCL) cell-of-origin (COO) distinction into
germinal center B cell (GCB) and activated B cell (ABC)
subtypes, as molecularly described by our group, has
profound biological, prognostic, and potential therapeutic
New therapeutic agents with selective activity in ABC and
GCB DLBCL are under development. An accurate diagnostic
assay is urgently needed to qualify patients for clinical
trials using targeted agents and as a predictive biomarker.
Although the subtypes were originally defined using gene
expression profiling on snap-frozen tissues (frozen-GEP), it
has become common practice to use less precise but
relatively inexpensive and broadly applicable
immunohistochemical (IHC) methods in formalin-fixed
paraffin-embedded tissue (FFPET).
We sought to create a robust, highly accurate molecular
assay for COO distinction using new GEP techniques
applicable to FFPET. This new assay shows excellent
performance in archival FFPET, and the rapid turn-around
time (<36 hours from FFPET block to result) will allow
prospective implementation in future therapeutic trials and,
ultimately, clinical practice.
Are We Ready To Stratify Treatment for Diffuse Large B-Cell
Lymphoma Using Molecular Hallmarks?
Current treatment paradigms
do not distinguish between COO phenotype, and this can be
readily justified. Several large analyses have shown no
difference in prognosis between phenotypes using IHC
classification methods, indicating a lack of precision and
reliability that dissuades against routine clinical use.
GEP has superior accuracy in classifying DLBCL into COO
subtypes with different biology and prognosis. It holds
promise for future clinical application once issues of
availability, cost, preference for analyses to be performed
on fresh frozen tissue, lack of standardization in COO
phenotype classification methods, and lack of prospective
randomized clinical evidence validating its use can be